title: A novel engineered anti-CD20 tracer enables early time PET imaging in a humanized transgenic mouse model of B-cell non-Hodgkins lymphoma.
abstract: OBJECTIVE
  The aim of this article was to evaluate the use of a novel engineered anti-CD20 protein based on the 10 kDa human fibronectin type 3 domain (FN3) and subsequently compare with (64)Cu-rituximab for positron emission tomography (PET) imaging of CD20.
  
  
  METHODS
  The engineered FN3(CD20) and FN3(WT) were produced in Escherichia coli cells at 2 to 5 mg/L, conjugated to DOTA, labeled with (64)Cu, and used for PET imaging of huCD20 expression in B cells. Humanized transgenic mice and subcutaneously xenografted mice each received intravenous (64)Cu-FN3(CD20) or FN3(WT) (3.7 MBq/4 μg Do-FN3 in 200 μL PBS). Control group received a blocking dose (50-fold excess) of unconjugated FN3(CD20) two hours before radiotracer injection. PET imaging was carried out at 1 to 24 hours postinjections.
  
  
  RESULTS
  In vitro assay demonstrated FN3 binds CD20 with 20 nmol/L affinity on CD20-expressing cells. (64)Cu-FN3(CD20) showed clear, high-contrast visualization of huCD20-expressing B cells in the spleen of transgenic mice as early as 1 hour postinjection [38 ± 3% injected dose (ID)/g] and exhibited a spleen-to-blood ratio of 13 by 4 hours. This is higher uptake (P = 0.04) and 10-fold greater signal-to-background (P = 0.04) than the (64)Cu-rituximab antibody radiotracer. Tumor uptake (16.8 ± 1.6 vs. 5.6 ± 1.4%ID/g) and tumor:background ratios were superior for FN3CD20 relative to rituximab in xenograft studies as well.
  
  
  CONCLUSIONS
  The (64)Cu-Do-FN3(CD20) radiotracer represents a novel small, high-affinity binder for imaging human CD20, which may be well suited for B-cell non-Hodgkin's lymphoma imaging in patients at early time points.
authors.affiliation: Authors' Affiliations: Molecular Imaging Program at Stanford (MIPS), Department of Radiology; and Bioengineering, Materials Science & Engineering, Stanford University, Stanford, California.
authors.forename: Arutselvan
authors.initials: A
authors.lastname: Natarajan
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Hackel
authors.forename: Sanjiv Sam
authors.initials: SS
authors.lastname: Gambhir
mesh_terms: D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000737:chemistry / Q000494:pharmacology; D018951:Antigens, CD20 / Q000737:chemistry* / Q000378:metabolism; D001402:B-Lymphocytes / Q000473:pathology; D003301:Copper Radioisotopes; D004195:Disease Models, Animal; D005353:Fibronectins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D016393:Lymphoma, B-Cell / Q000175:diagnosis* / Q000000981:diagnostic imaging / Q000473:pathology; D008228:Lymphoma, Non-Hodgkin / Q000175:diagnosis* / Q000000981:diagnostic imaging / Q000473:pathology; D051379:Mice; D008822:Mice, Transgenic; D049268:Positron-Emission Tomography; D011859:Radiography; D019275:Radiopharmaceuticals; D000069283:Rituximab; D014018:Tissue Distribution; D023041:Xenograft Model Antitumor Assays
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D018951:Antigens, CD20; D003301:Copper Radioisotopes; D005353:Fibronectins; D019275:Radiopharmaceuticals; C501364:fibronectin type III like peptide, human; D000069283:Rituximab